期刊
MOLECULAR THERAPY-NUCLEIC ACIDS
卷 32, 期 -, 页码 478-493出版社
CELL PRESS
DOI: 10.1016/j.omtn.2023.04.016
关键词
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APOBEC/AID cytidine deaminases play an important role in innate immunity and antiviral defenses. Researchers developed a CRISPR-activation-based approach to induce APOBEC/AID overexpression and control its effects on HBV replication and cellular toxicity. The study highlights the potential for precise control of APOBEC/AID activation as a strategy for suppressing HBV replication without toxicity.
APOBEC/AID cytidine deaminases play an important role in innate immunity and antiviral defenses and were shown to suppress hepatitis B virus (HBV) replication by deaminating and destroying the major form of HBV genome, covalently closed circular DNA (cccDNA), without toxicity to the infected cells. However, developing anti-HBV therapeutics based on APOBEC/AID is complicated by the lack of tools for activating and controlling their expression. Here, we developed a CRISPR-activation-based approach (CRISPRa) to induce APOBEC/AID transient overexpression (>4- 800,000-fold increase in mRNA levels). Using this new strategy, we were able to control APOBEC/AID expression and monitor their effects on HBV replication, mutation, and cellular toxicity. CRISPRa prominently reduced HBV replication (,,,90%-99% decline of viral intermediates), deaminated and destroyed cccDNA, but induced mutagenesis in cancer related genes. By coupling CRISPRa with attenuated sgRNA technology, we demonstrate that APOBEC/AID activation can be precisely controlled, eliminating off-site mutagenesis in virus-containing cells while preserving prominent antiviral activity. This study untangles the differences in the effects of physiologically expressed APOBEC/AID on HBV replication and cellular genome, provides insights into the molecular mechanisms of HBV cccDNA mutagenesis, repair, and degradation, and, finally, presents a strategy for a tunable control of APOBEC/AID expression and for suppressing HBV replication without toxicity.
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